BIOSENSORS & BIOELECTRONICS
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2021
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IF 10.618
Kai Zhang,Zhenqiang Fan,Bo Yao,Yuedi Ding,Jing Zhao,Minhao Xie,Jianbin Pan
Sialic acid-binding immunoglobulin (Ig)-like lectins (Siglecs) is a type I transmembrane receptor on the cell surface. Siglec-5, as one of the Siglecs family, play an important role as an inhibitory receptor for leukocytes in the human body. The development of novel siglec-5 assays can help to study the pathogenesis of related diseases as well as to develop novel therapeutic drugs. We use catalytic hairpin assembly (CHA) amplification strategy combined with CRISPR-Cas12a′s side-cutting feature to build a 2D ultra-thin Ti 3 C 2 T x (MXene) based electrochemiluminescence (ECL) biosensor for the detection of Siglec-5. By using this ECL biosensor, the cleavage of CRISPR-Cas12a is reasonably combined with CHA-mediated isothermal amplification, thereby realizing the sensitive amplification assay Siglec-5 with 20.22 fM sensitivity. By introducing pairs of sites that are not in the same double-stranded DNA into the DNA duplex, the hybridization sequence of CRISPR-Cas12a complements the targeting mechanism to enhance indirect Siglec-5 amplification assay. Also, the double-strand DNA (dsDNA) design based on CRISPR-Cas12a amplification allows the same CRISPR RNA (crRNA, also known as guide RNA (gRNA)) to detect the output of DNA duplexes from different intermediate DNAs, which provides a common way for biomarker detection based on the conversion of protein analytes to intermediate DNA strategy. This work extends the application scope of CRISPR-Cas12a to the construction of ECL biosensors, evaluates the role of lectins, which can be used for the biochemical research and clinical diagnosis of protein markers. This is the first investigative work exploring the Trans-Cleavage activity of CRISPR-Cas12a for Mxene-based ECL biosensor establishment to the best of our knowledge.
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高影响因子
Nano Today
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2025
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IF 10.9
Suleixin Yang,Ruie Chen,Peng Hua,Yi Wu,Meiwan Chen
Photothermal therapy (PTT) demands efficient cancer ablation at relative low temperatures and minimal thermal damage to normal tissues, but suffers from both the protective autophagy-related thermal resistance in cancer cell and reactive oxygen species (ROS)-induced damage to normal cells. Here, we screened out curcumin-Fe ultrasmall nanoparticles (Cur-Fe) that manifested efficient photothermal conversion efficiency (η = 43.38 %) and ROS scavenging ability. Additionally, CRISPR/Cas9 plasmids (pCas-ATG5/ATG7) were also constructed to safely and precisely knockdown the protective autophagy for thermal resistance alleviation. The core Cur-Fe/ATG@ TK PF (CFA@T), which was composed of anionic C ur- F e and pCas- A TGs, was encapsulated by cationic thioketal-crosslinked and fluorinated polyethyleneimine ( T K PF) via electrostatic interaction. Further, CFA@TC was formed by CFA@T coated with an acidic pH-responsive shell OH C -PEG-CHO via Schiff base. Attributed to its dual responsiveness to pH and ROS, CFA@TC exhibited efficient tumor targeting and uptake following intravenous injection. Upon irradiation with a 652 nm laser, CFA@TC demonstrated enhanced efficacy in eradicating cancer cells by inhibiting autophagy, while concurrently mitigating inflammatory responses through intracellular ROS scavenging in vivo and in vitro . Taken together, our study provides a proof-of-concept that CRISPR can be effective for autophagy inhibition, and its integration with ROS-induced inflammatory responses relief can further improve PTT.
研究领域:
癌症治疗
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纳米技术
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分子生物学
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生物医学工程
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高影响因子
LAB ON A CHIP
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2025
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IF 5.4
Zihan Wang,Fuwan Yang,Shuai Zeng,Rui Sun,Qifei Hu,Yichen Du
Applying CRISPR-based diagnostics to point-of-care pathogen detection remains challenging because of the multi-step and time-consuming sample preparation process. This study presents a low-cost, integrated valved microfluidic device that combines recombinase polymerase amplification (RPA), CRISPR signal amplification, and lateral flow readout for simultaneous nucleic acid detection. The core advantage of the platform lies in its ability to sequentially control the entire multi-step assay through simple valve operation, significantly minimizing user intervention. All key reagents, including the RPA mix, Cas12a/crRNA complex, and proteinase K lysis buffer, are pre-lyophilized, ensuring stability and ready-to-use functionality. The platform demonstrates a sensitivity of 20 copies/reaction for HPV16/18 plasmids and accurately genotypes HPV in lysates of cervical cancer cells within one hour, showing complete concordance with quantitative PCR results. This integrated device, achieving a user-friendly protocol and visual readout, provides a powerful tool for nucleic acid-based point-of-care testing and self-testing in resource-limited settings.
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TOXICOLOGY
·
2025
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IF 4.6
Zhisen Zhuang,Lijuan You,Jianbo Kong,Yaru Tian,Yongning Li,Yuan Zhi,Bo Zhang,Yi Wan,Xudong Jia,Hui Yang
Phthalate esters (PAEs) and per- and polyfluoroalkyl substances (PFAS) are ubiquitous pollutants tied to metabolic and immune disorders. The peroxisome proliferator-activated receptor (PPAR) pathway has been indicated to mediate their toxic effects, but the specific functions of PPAR subtypes and their mediating roles remain unclear. In this study, we generated PPARα-, δ-, and γ-specific knockout THP-1 cell lines by using CRISPR/Cas9 system and then differentiated them into interleukin-4 (IL-4) and interleukin-13 (IL-13)-polarized macrophages (alternative activation). During the induction process, the cells were exposed to 0, 6.25, 12.5, 25, 50, 100 μM of five PAE metabolites: Mono-(2-ethylhexyl) phthalate (MEHP), Monocyclohexyl phthalate (MCHP), Monoisonyl phthalate (MINP), Monoisobutyl phthalate (MIBP), and Monobenzyl phthalate (MBzP); and five PFAS: Perfluoroundecanoic acid (PFUnDA), Perfluorodecanoic acid (PFDA), Perfluorooctanoic acid (PFOA), Potassium perfluorooctanesulfonate (PFOS-K), and Potassium 9-chlorohexadecafluoro-3-oxanonane-1-sulfonate (F53B) for 48 h. The results showed that PPARδ deletion abolished CD209 expression, confirming its essential role, whereas PPARα deletion reduced and PPARγ deletion enhanced CD209, indicating PPARα promotes and PPARγ restrains alternative activation. Compounds displayed subtype-selective actions: MEHP activated PPARα/γ; MBzP/MCHP inhibited PPARδ yet activated PPARγ; MINP activated PPARγ only. Among PFAS, PFOS-K activated PPARδ/γ; F53B inhibited PPARα; PFOA activated PPARγ; PFDA inhibited PPARα/δ. Transcriptomics revealed compound-specific enrichments—cholesterol (MEHP), fatty-acid (MCHP), glycolysis (PFOS-K), TCA cycle (PFOA)—despite common PPAR pathway engagement. In conclusion, PAEs and PFAS disrupt macrophage plasticity through distinct PPAR-subtype signatures, providing molecular landmarks for future hazard assessment of environmental pollutants.
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Advanced Science
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2025
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IF 14.1
Yating Li,Jason Tan,Yanqin Tu,Jingnan Wu,Zaifang Zhang,Yifeng Wei,Xinan Jiao,Yan Zhou
DNA base modification is a common strategy used by bacteriophages to evade host immune detection. A prominent example is dU-DNA, where thymidine is globally replaced with 2′-deoxyuridine. Despite its widespread occurrence, dU-DNA's biosynthetic pathways and functional roles remain incompletely understood. Here, different enzymes supporting dU-DNA biosynthesis in phage PBS1, Roseophage DSS3_VP1, and Yersiniophage PhiR1-37 are identified and characterized. The dU nucleotide precursor is supplied by phage dCTP deaminases (Dcds). Thymidine nucleotides are degraded by dTMP phosphatases (Dtms) in PBS1 and DSS3_VP1, and by a dTTP pyrophosphatase (Dtt) in PhiR1-37, preventing incorporation into phage DNA. The dU-DNA, isolated from Yersiniophage PhiR1-37 or synthesized by PCR, demonstrates resistance to cleavage by restriction enzymes recognizing thymidine-containing sequences, partial resistance to Lb Cas12a nuclease recognizing a TTTV PAM site, while remaining sensitive to Sp Cas9 nuclease recognizing a NGG PAM site. A phylogenetic analysis of PBS1 dCTP deaminase and closely related T4 phage dCMP deaminase suggest possible evolutionary origins from bacterial dCDP deaminases. Overall, these findings suggest independent acquisition of dU-DNA biosynthetic enzymes and pathways in the diverse phages, and support its protective function against different host-encoded nucleases.
研究领域:
分子生物学
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病毒学
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微生物学
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基因工程
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核酸代谢
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免疫逃避
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进化生物学
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高影响因子
Biomaterials Science
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2022
·Vol. 10
·期 21
·6230-6243
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IF 7.59
Yuanji Feng,Zhaopei Guo,Jie Chen,Sijia Zhang,Jiayan Wu,Huayu Tian,Xuesi Chen
To improve the efficiency of nucleic acid and protein delivery by cationic polymers, there is a trade-off between increasing the positive charge density of cationic polymers and decreasing cytotoxicity. In this work, a strategy to introduce multiple interactions between the cell membrane and a delivery system based on cationic polymers was proposed. A novel delivery system consisting of PEI1.8k and an enhancer (LA-RT) was fabricated. The introduction of LA-RT contributed to multiple interactions between the delivery system and the cell membrane including electrostatic interactions, hydrogen bonding, hydrophobic interaction, and dynamic sulfur exchange reactions, which enabled efficient intracellular delivery of nucleic acids and proteins. For nucleic acid delivery, plasmid DNA and mRNA were loaded to realize CRISPR/Cas 9 gene editing in vivo and protein expression in vivo, respectively. For protein delivery, the delivery system carrying OVA protein and CpG formed a nano-vaccine, which induced enhanced humoral and cellular immunity in vivo. In addition, the delivery system based on PEI1.8k revealed negligible cytotoxicity. This work provided a novel strategy to prepare efficient delivery systems based on cationic polymers via the introduction of a multifunctional enhancer.
研究领域:
药学
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分子生物学
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生物技术
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免疫学
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高影响因子
Frontiers in Bioengineering and Biotechnology
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2024
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IF 4.3
Huang Hao,Li Shuang,Han Xianjing,Zhang Yule,Gao Lingfeng,Wang Xiangjiang,Wang Guiqing,Chen Zhi
Osteoarthritis (OA) has become a serious problem to the human society for years due to its high economic burden, disability, pain, and severe impact on the patient’s lifestyle. The importance of current clinical imaging modalities in the assessment of the onset and progression of OA is well recognized by clinicians, but these modalities can only detect OA in the II stage with significant structural deterioration and clinical symptoms. Blood vessel formation induced by vascular endothelial growth factor (VEGF) occurs in the early stage and throughout the entire course of OA, enables VEGF relating gene sequence to act as a biomarker in the field of early diagnosis and monitoring of the disease. Here in, a facile rapid detection of VEGF relating ssDNA sequence was developed, in which manganese-based zeolitic imidazolate framework nanoparticles (Mn-ZIF-NPs) were synthesized by a simple coprecipitation strategy, followed by the introduction and surficial absorption of probe ssDNAs and the CRISPR/Cas12a system components. Furthermore, fluorescence experiments demonstrated that the biosensor displayed a low detection limit of 2.49 nM, a good linear response to the target ssDNA ranging from 10 nM to 500 nM, and the ability of distinguishing single nucleotide polymorphism. This finding opens a new window for the feasible and rapid detection of ssDNA molecules for the early diagnose of OA.
研究领域:
生物医学工程
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分子生物学
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纳米技术
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诊断学
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高影响因子
MICROCHEMICAL JOURNAL
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2025
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IF 5.1
Weiying Liu,Xiaolu Sui,Yunpeng Xu,Tingfei Xie,Yanzi Zhang,Aisha Zhang,Zhenyu Lin,Jihong Chen
MicroRNA-21 (miRNA-21) can serve as an early biomarker for acute kidney injury (AKI), it is important to develop sensitive and selective detection methods for miRNA-21 detection. In this study, an electrochemiluminescence (ECL) biosensor for miRNA-21which synergistically integrates T7 RNA polymerase-driven transcriptional amplification with CRISPR/Cas13a-mediated collateral cleavage and magnetic separation strategy had been designed. T7 RNA polymerase-driven transcriptional amplification has the characters of low requirements for conditions and high amplification efficiency. A cascaded signal amplification strategy which coupled enzymatic target recycling with CRISPR/Cas13a-mediated exponential signal amplification was proposed to achieve high detection sensitivity. And T7 promoter specificity and CRISPR/Cas13a sequence targeting had been adopted to ensure high specificity of the developed system. Furthermore, magnetic bead separation had been adopted to reduce the background signal. The parameters which may affected the performance of the biosensor had been optimized. Under optimized conditions, the ECL response of the system has a linear relationship with the logarithmic value of miRNA-21 concentration in the range of 1.0 fM to 1.0 nM, with a calculated limit of detection(LOD) of 0.33 fM (S/N = 3). The proposed methodology has been adopted to detect the target in clinical urine specimens with satisfied results.
研究领域:
生物化学
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分子生物学
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分析化学
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生物医学工程
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临床诊断
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高影响因子
JOURNAL OF AGRICULTURAL AND FOOD CHEMISTRY
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2023
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IF 6.1
Xiaobo Zhang,Zhiru Li,Lili Yang,Bing Hu,Qiuyue Zheng,Jiang Man,Jijuan Cao
The sensitive and accurate detection of ochratoxin A (OTA) is crucial for public health due to its high toxicity. Herein, using Au nanoparticle (NP)-attached CdS/UiO-66-NH2 heterostructures as photoactive materials, a photoelectrochemical (PEC) aptasensor was presented for the ultrasensitive assay of OTA based on a competitive displacement reaction triggering the trans-cleavage ability of CRISPR/Cas12a. In this sensing strategy, methylene blue-labeled single-stranded DNA (MB-ssDNA) was immobilized on the Au NPs/CdS/UiO-66-NH2 electrode to accelerate the separation of the photogenerated carrier, thus producing a significantly increased PEC response. In the presence of OTA, it specifically bound with the aptamer (Apt) and resulted in the release of the activation chain, triggering the trans-cleavage characteristics of CRISPR/Cas12a. MB-ssDNA was cut randomly on the electrode surface to convert the PEC signal from the “on” to the “off” state, thereby achieving a quantitative and accurate detection of OTA. The CRISPR/Cas12a-derived PEC aptasensor exhibited excellent sensitivity and specificity, with a linear range from 100 to 50 ng/mL and a detection limit of 38 fg/mL. Overall, the proposed aptasensor could provide a rapid, accurate, and sensitive method for the determination of OTA in actual samples.
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SENSORS AND ACTUATORS B-CHEMICAL
·
2023
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IF 8.4
Pei Wang,Bohui Zhou,Ziye Liu,Yining Zhao,Robert Chunhua Zhao,Jinhui Peng,Juan Zhang
The accurate detection of immune checkpoint molecules on the surface of tumor cells is of great significance for immunotherapy. Herein, we propose a method based on cellular interface supported cascade signal amplification mediated by CRISPR/Cas13a system for in situ analysis of immune checkpoint molecules on bladder cancer cells. CD47 and EpCAM on bladder cancer cells can be sensitively monitored through T7 transcription amplification, CRISPR/Cas13a system and HCR triple signal amplification. Moreover, the established method can specifically differentiate EpCAM and CD47 positive bladder cancer cells and well serve for the evaluation of two kinds of membrane protein inhibitors. In a word, the method we proposed can successfully realize in situ analysis of immune checkpoint molecules on bladder cancer cells for guiding immunotherapy.
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Journal of Materials Chemistry B
·
2022
·Vol. 10
·期 32
·6107-6117
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IF 7.571
Siying Xie,Cheng Qin,Furong Zhao,Zhaoyang Shang,Pei Wang,Muhammad Sohail,Xing Zhang,Bingzhi Li
CRISPR-driven biosensing is developing rapidly, but current studies mostly adopt dye-labeled ssDNA as the signal reporter, which is costly and unstable. Herein, we developed a label-free and low-background reporter for CRISPR/Cas12a signaling by integrating DNA-templated copper nanoclusters (DNA–CuNCs) and exonuclease I (EXO I). The template of the DNA–CuNCs was rationally designed as a ds-/ss-DNA hybrid, ensuring that after a quick and nonpersistent cut of Cas12a, a majority of the template can be digested by EXO I. Based on this novel reporter, a biosensor termed CRISPR-CNS (cost-effective, nimble, and sensitive copper nanocluster sensor integrating CRISPR) was developed. Due to the high signal-to-background ratio of our proposed reporter, CRISPR-CNS shows excellent performances for nucleic acid detection, yielding a detection limit of 20 copies for SARS-CoV-2 RNA. Considering its facile synthesis, robust fluorescence, effective cost, and good sensitivity, this combination shall serve as a highly potential output for CRISPR-based point-of-care testing.
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SENSORS AND ACTUATORS B-CHEMICAL
·
2022
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IF 9.221
Li Shuofeng,Wang Fangfang,Chun Wang,Zhi Wang,Qiuhua Wu
As vital biomarkers for diagnosis of diseases, the anomalously expressed microRNAs (miRNAs) are closely linked to the occurrence and development of many diseases. Quantitative analysis of miRNAs has remarkable significance to clinical diagnosis and therapy of different cancers. In this study, the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR-associated (Cas) 13a (CRISPR/Cas13a) system was first exploited to directly and specifically recognize the miRNA at the guidance of CRISPR RNA (crRNA), which can activate the nonspecific trans-ribonuclease activity of Cas13a. Then, combining the CRISPR-Cas13a system with terminal deoxynucleotidyl transferase (TdT)-triggered template-free DNA extension to form the fluorescent copper nanoparticles (CuNCs), a simple but sensitive method (named as Cas13a-TFCN) was developed for highly specific detection of miRNA. Based on the CRISPR/Cas13a system coupled with TdT-triggered simple but efficient signal amplification strategy, the developed Cas13a-TFCN method showed high sensitivity for miRNA detection with the low detection limit of 0.7 fM. Furthermore, the proposed Cas13a-TFCN approach was successfully applied for accurate quantification of miRNA in complex biological samples, indicating its practical application ability. Moreover, featuring high sensitivity and simplicity without complicated probe design and fluorophore labeling, the proposed Cas13a-TFCN has a promising applications prospect in clinical diagnosis.
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